We have determined that gustducin, T1R3 and several other signaling elements that underlie taste detection/transduction also are expressed in L and K subtypes of enteroendocrine cells. One function of gustducin and T1R3 in L cells is to sense glucose or sweeteners within the lumen of the gut. 1-Gustducin knockout mice do not secrete GLP-1 from their enteroendocrine L cells in response to infusion of glucose into the gut lumen. Because of this defect these mice have dysfunctional regulation of their plasma insulin and glucose levels. Studies with minced duodenal tissue or isolated duodenal villi from 1-gustducin knockout mice show that the defect in GLP-1 secretion is independent of innervation. Studies with human and mouse enteroendocrine cell lines and antisense blockade of gustducin or pharmacological inhibition of T1R3 indicate that both gustducin and T1R3 are required for enteroendocrine cell release of GLP-1, suggesting that the defect in 1-gustducin knockout mice in vivo is at the level of glucose sensation. The experiments proposed here aim to further characterize the functions of gustducin, T1R3 and other taste signaling elements in enteroendocrine cells. We have combined molecular, cellular, transgenic, physiological and pharmacological methods to: 1. Determine which key taste signaling molecules are expressed in which enteroendocrine cells; 2. Determine if hormone release from enteroendocrine cells depends on their expression of taste signaling elements and elicits upregulation of sugar and fatty acid transporters in enterocytes; 3. Determine if pharmacologic or genetic block of enteroendocrine cell-expressed taste signaling elements alters hormone release from enteroendocrine cells; 4. Generate conditional knockout mice selectively lacking 1-gustducin, T1r3 or Trpm5 in gut; 5. Determine if conditional knockout mice lacking 1- gustducin, T1r3 or Trpm5 in gut are resistant to diet-induced obesity, show defective 2 cell function on a high fat diet, and exhibit increased energy expenditure. We will be testing the following key hypotheses: 1. Specific types of enteroendocrine cell type express multiple taste signaling elements; 2. Macronutrients and tastants elicit release of GLP-1 and other hormones from enteroendocrine cell by activating gustducin and taste receptors expressed in subtypes of enteroendocrine cells; 3. Conditional Knockout mice lacking enteroendocrine cell-expressed taste signaling elements will have defects in regulating their plasma levels of GLP-1, GIP and other hormones - leading to: (a) failure to upregulate certain enterocyte transporters, (b) dysregulation of glucose homeostasis, (c) resistance to diet-induced obesity, (d) disruption of 2 cell functions, (e) increased energy expenditure. This multidisciplinary approach has promise for providing significant new insights into the nature of enteroendocrine cell function. In particular, how enteroendocrine cells sense macronutrients (e.g. glucose and sugars) and tastants (e.g. artificial sweeteners) in the gut lumen, and how this leads to stimulation of release of GLP-1 and other hormones to regulate gut functions. This proposal has medical relevance to appetite, satiety, obesity, and diabetes.